Cardiovascular and cerebrovascular diseases are the leading causes of death worldwide. Obesity, elevations in circulating levels of cholesterol and triglycerides and alterations in the composition of circulating lipoproteins are major risk factors for cardiovascular and cerebrovascular disease. Obesity is a disorder of energy balance. When energy input exceeds energy output most of the excess calories consumed are converted to triglycerides and stored in adipose tissue. Similarly, increases in circulating cholesterol leading to cholesterol deposition in the vasculature occur when the balance between de novo synthesis of cholesterol and provision of cholesterol from dietary sources exceeds normal cholesterol requirements and capacity to eliminate this essential sterol. Clinical management of obesity and its complications including hypercholesterolemia presently focuses on modifications of diet, physical activity and behavior. Because these strategies are of limited effectiveness, the prevalence of obesity continues to increase. Pharmacological inhibition of cholesterol synthesis is the primary and presently most effective strategy for the clinical management of hypercholesterolemia. Although to date efforts to develop pharmacological strategies for treatment of obesity focusing on appetite suppression and interference with lipid absorption have met with moderate success, recent advances using mouse models suggest that inhibition of triglyceride synthesis may be a viable and effective treatment strategy. Identification of components of the pathways responsible for synthesis of triglycerides and cholesterol and development of a complete understanding of how these processes are regulated is therefore critical for the development of new or improved agents for treatment of obesity and hypercholesterolemia. Because of their pivotal position in cellular metabolism, the pathways responsible for the de novo synthesis of cholesterol and triglycerides have been intensively studied but significant gaps in our knowledge remain. Two classes of lipid phosphate esters, polyisoprenoid diphosphates and phosphatidic acid are critical intermediates in the synthesis of cholesterol and triglyceride but enzymes that dephosphorylate these intermediates have only recently been identified. The overall goal of this research is to address these gaps in our knowledge by investigating the regulation and function of two newly identified lipid phosphatases that play central roles in the synthesis of isoprenoids, sterols and triglycerides. Our research uses techniques of biochemistry, cell and molecular biology to study the regulation and function of these enzymes in vitro and in biomedically relevant model cell systems. In the first aim we will use these approaches to test the hypothesis that an integral membrane enzyme termed polyisoprenoid diphosphate phosphatase is a regulator of cholesterol synthesis and protein isoprenylation. In the second aim of the proposal we will test the hypothesis that activity of a phosphatidic acid phosphatase enzyme that catalyses a critical step in triglyceride synthesis is regulated by a novel membrane targeting motif. PUBLIC HEALTH RELEVANCE: Obesity and elevated cholesterol levels are major risk factors for Cardiovascular and Cerebrovascular disease which are the leading causes of death in this country. Obesity is a disorder of energy balance. When energy input exceeds energy output most of the excess calories consumed are converted to triglycerides and stored as body fat. Similarly, increases in circulating cholesterol occur when the balance between cholesterol synthesis and provision of cholesterol from dietary sources exceeds normal cholesterol requirements and cholesterol excretion. In this research project we will test specific hypotheses about the regulation and function of newly identified enzymes that play central roles in the synthesis and metabolism of fat and cholesterol. Completion of our work will provide new insights into how synthesis of these lipids is regulated, how this process may be altered in disease and the feasibility of targeting these processes to provide novel therapies for obesity and cardiovascular disease. [unreadable] [unreadable] [unreadable]